Hyaluronic acid is a linear macromolecular polysaccharide consisting of alternately bonded β-D-N-acetylglucosamine and β-D-glucuronic acid. Hyaluronic acid is found not only in connective tissues of mammals but also in cockscombs and the capsules of Streptococci. Hyaluronic acid is obtainable not only by extraction from cockscombs and umbilical cords, but also as purified products from the culture broth of Streptococci.
Natural hyaluronic acid is polydisperse in respect of molecular weight and is known to show excellent biocompatibility even when implanted or injected into the body by virtue of the absence of species and organ specificity.
Synovial fluid supplies nutrition to the articular cartilage and has incomparable functions as a lubricant and a shock absorber in the joint, and its excellent viscoelasticity heavily owes to one of the main components in the synovial fluid, hyaluronic acid. Hyaluronic acid is known to have a high molecular weight of from several million to ten million.
Concentration and molecular weight analyses of hyaluronic acid in the synovial fluid from patients with arthropathy such as osteoarthritis and chronic articular rheumatism demonstrated the concentration and molecular weight of hyaluronic acid in the synovial fluid from patients with arthropathy generally tend to lower than in normal synovial fluid, and it is considered that the lower concentration and molecular weight of hyaluronic acid are closely associated with development of locomotor dysfunction and pain attributable to the weaker lubricating action and the weaker protecting action on the surface of the articular cartilage of synovial fluid.
Injection of a hyaluronic acid solution into diseased joints has been adopted as an effective measure for knee osteoarthritis among those articular diseases. As examples of a therapeutic agent for knee joints, Artz (manufactured by Seikagaku Corporation, average molecular weight 900,000), Hyalgan (manufactured by Fidia, average molecular weight <500,000), or Suvenyl (manufactured by Aventis Pharma Ltd., CHUGAI PHARMACEUTICAL CO., LTD. and DENKI KAGAKU KOGYO KABUSHIKI KAISHA, average molecular weigh 1,900,000) which has been developed by the present inventors, with the idea that a higher effect can be expected with a higher molecular weight, which is also adapted to chronic articular rheumatism, may be mentioned.
Further, Synvisc (Genzyme) containing a crosslinked hyaluronic acid gel, in which hyaluronic acid is chemically crosslinked to make it have a high molecular weight, thereby to improve the viscoelasticity, has been developed. The crosslinked hyaluronic acid gel is a hyaluronic acid gel formed by chemically crosslinking hyaluronic acid with a crosslinker divinyl sulfone and is called Hylan. Methods for producing Hylan and a gel obtained by crosslinking Hylan are disclosed in U.S. Pat. No. 4,713,448 in detail. Single and mixed gels comprising hyaluronic acid as the base are disclosed in U.S. Pat. No. 4,582,865 and U.S. Pat. No. 4,605,691.
For treatment employing such preparations for joints, direct injection to the joints of patients 3 to 5 times every week is required, and decrease in the number of injections has been desired in view of the burden on the patients and doctors.
A therapeutic agent for joints containing self-crosslinking hyaluronic acid is also disclosed in WO97/49412. The object is to control viscoelastic properties and to prolong the residence time in the joint by crosslinking hyaluronic acid. A technique to control the viscoelasticity by uniformly mixing self-crosslinking hyaluronic acid and an aqueous solution of hyaluronic acid is disclosed. It is also conceptually disclosed that self-crosslinking hyaluronic acid is hydrolyzed in the articular cavity and can become natural hyaluronic acid containers (reservoirs).
The pH of the synovial fluid is from about 7.3 to about 7.6. It is pointed out that a pH of a therapeutic agent for joints to be injected into the articular cavity which is significantly different from the pH of the synovial fluid may cause irritation or pain (SEIKAGAKU DATA BOOK I, THE JAPANESE BIOCHEMICAL SOCIETY, TOKYO KAGAKU DOZIN CO., LTD., 1979, p 1569). Accordingly, the pH of Artz and Suvenyl is adjusted to be within a range centering on 7.4.
The form of distribution has been greatly changed from vial preparations to kit preparations such as prefilled syringes, also with respect to the therapeutic agent for joints, for the purpose of labor-saving at the time of preparation for administration at a medical institution and preventing bacterial contamination and contamination by foreign substances. It takes long to fill a syringe with a particularly highly viscous hyaluronic acid preparation by suction, and the amount in the syringe significantly fluctuates. Kit preparations are shown to be a medically effective form, with which the burden of preparation for administration at a medical institution is reduced and at the same time, a set amount of the liquid can be easily and more uniformly injected.
In the process of extensive studies on properties of a hyaluronic acid ester derivative, particularly self-crosslinking hyaluronic acid, the present inventors have found that the ester derivative has no adequate stability in a neutral region. A medical material to be administered to the body is prepared to be in a neutral region as a biological condition, however, a possibility is considered that when a hyaluronic acid ester derivative is adapted as a medical material under such a condition, it can not be stored for a long time in the process of e.g. distribution.
Accordingly, particularly for preparation of a medical material comprising such a hyaluronic acid ester derivative, in order to overcome such a problem that the hyaluronic acid ester derivative is unstable in a neutral pH region, the present inventors have been conducted extensive studies on a method for stably providing such an ester derivative. As a result, they have found that the ester derivative has a higher stability under a weakly acidic condition in a pH region of from 4.5 to 6.5. Further, they have conceived, to provide a medical material comprising a hyaluronic acid ester derivative as a component, such a preparation that a dispersed aqueous solution of a weakly acidic hyaluronic acid ester derivative and a neutralizing agent solution therefor, including a case where hyaluronic acid is dissolved in the neutralizing agent solution, are packed separately from each other so that they are under different pH conditions, they can be mixed for injection at the time of treatment, and finally they can be administered under a physiologic neutral pH condition, and they have accomplished the present invention.